Purpose: Glioblastoma (GBM) are aggressive astrocytomas typified by progressive diffuse infiltration and surgical complications. There is an urgent need to elucidate and target factors that drive tumour progression. The expression of CD147 (a matrix metalloproteinase (MMPs) inducer and the receptor for cyclophilin A (CypA) mediated signalling) is correlated with GBM severity. CypA is a potent chemokine and cytoprotectant associated with GBM tumour severity through CD147 mediated MMP9 expression. Microglia are an essential component of the tumour microenvironment and support tumour growth via cytokine secretion. Thus we aim to determine if microglia can secrete CypA and determine if extracellular CypA can support GBM pathogenesis.
Methods: BV2 microglia were treated with lipopolysaccharide (LPS) and the oxidative stress inducer LY83583 at a range of time points, and their supernatants subjected to WB analysis to determine the presence of secreted CypA protein. BV2 microglia were also treated with recombinant CypA (rCypA) protein (1nM-1000nM) to assess changes in cell migration and cell proliferation. U251 and U87 GBM cell lysates, treated with rCypA protein (100nM), were subjected to WB analysis to assess ERK 1/2 and AKT phosphorylation.U251 and U87 cells were pre-treated with rCypA (10 and 100nM) for 1h followed by cisplatin for up to 72h. LDH and MTS assays were used to determine cell viability.
Results: We found that BV2 microglia secreted CypA in response to LPS and LY83583. Furthermore, rCypA protein increased cell migration at 24h and reduced basal cell death after 72h in BV2 microglia We also demonstrated that rCypA protein induces ERK 1/2 and Akt phosphorylation in the U251 and U87 GBM cells. Furthermore, 1hr rCypA pre-treatment reduced cisplatin induced cell death in both GBM cell lines.
Conclusion: We have shown that microglia secrete CypA and that CypA can activate cell surivival signalling and drive chemoresistance to cisplatin in GBM cells.